Human lutropin or human luteinizing hormone (hLH) belongs to the glycoprotein hormone family along with human follicle stimulating hormone (FSH), human thyroid stimulating hormone (TSH) and human chorionic gonadotropin (hCG) (for review see Ryan R. J. et al. 1987, Structure-function relationships of gonadotropins. Rec Prog Horm Res 43:383-429). The glycoprotein hormones consist of two subunits. The amino acid sequence of the alpha subunit is identical or nearly identical for the four human glycoprotein hormones. Despite significant homologies between the different glycoprotein hormones, the beta subunit is responsible for the target organ specificity of the hormone. hLH and hFSH which are produced by cells of the pituitary gland, regulates the steroid hormone production in the human gonads. The beta subunits of hLH and hCG are highly homologous. The major difference is the additional 24 amino acid extension at the carboxyterminal of hCG.beta. subunit. The biological action of hLH and hCG is mediated through the same receptor.
The carbohydrate part plays a significant role in the biological action of the glycoprotein hormones. Chemical deglycosylation has been used to study the function of Asn linked oligosacharide moieties in glycoprotein hormone action. This procedure does not affect the interaction between the alpha and beta dimers nor the binding of the hormone to its receptor. The deglycosylated hormones, however, have significantly reduced ability to stimulate the cAMP production in target cells despite normal binding to the receptor. The biological effect of the hormone is also indirectly influenced by the carbohydrate part, as differences in the structure of the carbohydrate moieties will effect the circulatory half lives of the hormone (Baenziger J. U. et al. 1992. Circulatory half-life but not interaction with the lutropin/chorionic gonadotropin receptor is modulated by sulfation of bovine lutropin oligosacharides. Proc Natl Acad Sci USA 89:334-338). hLH and hCG, although highly similar in peptide sequence and in vitro biological activity, have quite different in vivo biological half lives. The oligosacharides of hLH when synthesized in the gonadotrophs are largely sulfated through mediation of enzyme systems specific for the gonadotroph cells of the pituitary. Increased sulfation gives hLH a shorter circulatory half-life due to removal by hepatic cell receptors specific for the sulfated forms of LH (Fiete D. et al. 1991. A hepatic reticuloendothelial cell receptor specific for SO.sub.4 -4GalNAc.beta.1, 4GlcNAc.beta.1, 2Man-alpha that mediates rapid clearance of lutropin. Cell 67:1103-1110). The carbohydrates of hCG synthesized in trophoblastic cells are not sulfated and therefore the protein has a longer circulatory half-life. Interspecies differences in glycosylation of the same glycoprotein as well as differences between the different glycoprotein hormones are obviously determined partly by differences in peptide sequence and partly by the glycosylation enzymes of the hormone producing cells (for review see Green E. D. et al. 1986. Differential processing of Asn-linked oligosaccharides on pituitary hormones: implications for biologic function. Mol Cell Biochem 72:81-100).
Due to differences in the carbohydrate part of the molecules, the glycoprotein hormones display a multitude of isoforms as seen in separating systems based on charge (Wide L. 1985. Median charge and charge heterogeneity of human pituitary FSH, LH and TSH. I. Zone electrophoresis in agarose suspension. Acta Endocrinol 109:181-189).
The concentration of hormones in biological fluids can be estimated through in vivoor in vitro biological assay systems. However, determinations with immunoassays provide a more convenient way for estimation of hormone concentrations/activites. Differently designed immunoassays constitute the preferred methods of hormone determinations in routine clinical laboratories. With monoclonal antibodies, sensitive detection technologies and non-competitive assay designs, extremely specific and sensitive assay sytems have been optimized.
In a previous work (Pettersson K. S. I et al. 1991, Individual differences in LH immunoreactivity revealed by monoclonal antibodies. Clin Chem 37:333-340) on optimizing non-competitive (sandwich) immunoassays for hLH employing monoclonal antibodies we found that certain monoclonal antibodies recognizing epitopes present on the intact alpha-beta dimer (but not on the free subunits) of LH displayed restrictive reactivity against hLH in some individuals. Depending on the monoclonal antibodies used, the LH immunoactivity in some individuals were either reduced to various degrees or altogether non-detectable. A pedigree study (Pettersson K et al. An immunologically anomalous LH variant in a healthy woman. J Clin Endocrinol Metab 74:164-171, 1992) of the proband initially found (a 31 year old woman with no detectable hLH when employing a certain monoclonal antibody) strongly indicated that these observations were due to a genetic variant of hLH (variant LH [V-LH]) as opposed to normal LH [N-LH]. Measurement of in vitro LH biological activity also showed that hLH of these individuals with immunologically "silent" LH employing certain monoclonal antibodies does possess normal biological activity (Pettersson K et al. An immunologically anomalous LH variant in a healthy woman. J Clin Endocrinol Metab 74:164-171, 1992; Pettersson K et al. 1991, Monoclonal antibody-based discrepancies between two-site immunometric tests for lutropin. Clin Chem 37:1745-1748). Our studies also indicate that the putative heterozygous condition (individuals with both N-LH and VLH) is quite frequent--about 20-25 percent in Finnish and Scandinavian populations (Pettersson K. S. I et al. 1991. Individual differences in LH immunoreactivity revealed by monoclonal antibodies. Clin Chem 37:333-340).